Mapping genomic regions associated with Maize Lethal Necrosis (MLN) using QTL-seq
Six bi-parental populations were
evaluated in three seasons under
artificial inoculation using a combination
of MCMV and SCMV (Fig. 1) and
genotyped with 156 to 289 polymorphic
SNPs using the KASP assay (Semagn
et al. 2014) and GBS (Elshire et al.
Five additional F2 mapping populations
were developed, genotyped with DART
SNPs and phenotyped under artificial
Three major QTLs on chromosomes 3
and 6 and a few minor QTL across other
chromosomes were identified (Fig. 2)
Major MLN disease resistant QTL on
chromosome 6 was validated by QTL
seq and confirmed (Fig. 3)
Mapping genomic regions associated with Maize Lethal
Necrosis (MLN) using QTL-seq
1*Mike Olsen, 2*Nasser Yao, 1Berhanu Tadesse, 1Bish Das, 1Manje Gowda, 1Kassa Semagn, 1MacDonald Jumbo, 3Andrzej Killian
1International Maize and Wheat Improvement Center (CIMMYT); 3Diversity Array Technology (DArT), Canberra, Australia
2Biosciences eastern and central Africa-International Livestock Research Institute (BecA-ILRI) Hub, Nairobi, Kenya
*For more information contact Nasser Yao/Mike Olsen
Partner institutions: International Maize and Wheat Improvement Center (CIMMYT),
Nairobi, Kenya; Diversity Arrays Technology Pty Ltd (DArT), Canberra, Australia
Funding: Bill and Melinda Gates Foundation (BMGF)
This document is licensed for use under a Creative Commons Attribution –Non commercial-Share Alike 3.0 Unported License February 2016
Maize lethal necrosis (MLN) is caused by the co-infection of maize with maize chlorotic mottle virus (MCMV) and any of the cereal viruses in the
Potyviridae family, including sugarcane mosaic virus (SCMV), maize dwarf mosaic virus or wheat streak mosaic virus (Cabanas et al., 2013). SCMV
was reported in Kenya over three decades ago (Louie, 1980) and is known to cause a minor losses. The introduction of MCMV in Kenya (Wangai et
al., 2012) and its ability to co-infect with SCMV leads to the development of MLN, posing a serious threat to maize productivity and livelihoods of the
farming communities in eastern Africa. The first sign of MLN was reported in Kenya in September 2011 near Bomet. There is an urgent need to
identify MLN resistance source germplasm, map the genomic regions associated with MLN resistance, and introgress MLN resistance loci from
suitable donors into the genetic backgrounds of widely used inbred lines and hybrids.
CIMMYT has undertaken discovery studies to identify putative genomic regions associated with MLN disease resistance. Two association mapping
panels and six bi-parental populations were evaluated for MLN disease severity under artificial inoculation (Gowda et al. 2015). All lines were
genotyped with 156 to 289 polymorphic SNPs using the Kompetitive Allele Specific PCR (KASP) assay (Semagn et al., 2014) and genotyping by
sequencing (GBS, Elshire et al., 2011). The analysis of the data from the bi-parental populations revealed three major QTL on chromosomes 3 and
6, and a few minor QTL across other chromosomes.
Currently, CIMMYT is working to introgress MLN resistance into adapted germplasm using both conventional backcrossing and marker-assisted
backcrossing (MABC). Based on results from the QTL mapping studies, it is currently being introgressed few major genomic regions using 7 MLN
donor lines to improve MLN tolerance level across 16 highly popular but MLN susceptible inbred lines widely grown in Africa. Based on the result of
the initial QTL mapping studies, it became necessary to conduct additional MLN QTL mapping studies using bulk segregant analysis and QTL
sequencing, which has been proposed as a rapid method for QTL detection (Lu et al., 2014; Takagi et al., 2013). For this purpose, five F2 mapping
populations were developed and phenotyped under artificial inoculation. Based on the MLN disease score, the most resistant and susceptible
progenies were selected for QTL seq, each population represented by 31-71 resistant and 29-67 susceptible progenies.
The overall aim on the project was to: (i) deliver improved versions of the elite African lines having 0.6 to 1.0 point lower MLN severity score and
contributing at least 10% higher grain yield (GY) advantage to hybrid combinations under MLN disease pressure as compared to the near isogenic
hybrids involving the recurrent parent; and (ii) having less than 5% GY reduction in near isogenic hybrid combinations in the absence of MLN disease
QTLs associated with MLN resistance
through whole genome sequencing of
MLN contrasting phenotypes tagged,
mapped and characterized
Improved elite African lines having MLN
tolerance and associated QTLs
identified and used for marker assisted
Improved versions of the elite African
lines having 0.6 to 1.0 point lower MLN
severity score and contributing at least
10% higher grain yield advantage to
hybrid combinations under MLN
disease pressure are available.
Less than 5% grain yield reduction in
near isogenic hybrid combinations in
the presence of MLN pressure are
• Whole genome sequencing of both resistant and susceptible bulk
population along with their parents
• Development of Maize Lethal Necrosis resistant lines that will be used in
hybrid breeding program and ultimately released to farmers
Figure 3: A major MLN QTL detected and validated on chromosome 6 association
Figure 2: Major and minor MLN QTLs detected across chromosomes 3, 6, 8 and 10
Cabanas D., Watanabe S., Higashi C.H.V., Bressan A. (2013) Dissecting the mode of maize chlorotic mottle virus transmission (Tombusviridae: Machlomovirus) by Frankliniella williamsi (Thysanoptera: Thripidae). Journal of Economic Entomology 106:16-24. DOI: 10.1603/ec12056.
Elshire R.J., Glaubitz J.C., Sun Q., Poland J.A., Kawamoto K., Buckler E.S., Mitchell S.E. (2011) A robust, simple genotyping-by-sequencing (GBS) approach for high diversity species. PLoS ONE 6:e19379.
Lu H., Lin T., Klein J., Wang S., Qi J., Zhou Q., Sun J., Zhang Z., Weng Y.Q., Huang S. (2014) QTL-seq identifies an early flowering QTL located near Flowering Locus T in cucumber. TAG Theoretical and Applied Genetics 127:1491-1499.
Semagn K., Babu R., Hearne S., Olsen M. (2014) Single nucleotide polymorphism genotyping using Kompetitive Allele Specific PCR (KASP): overview of the technology and its application in crop improvement. Molecular Breeding 33:1-14. DOI: 10.1007/s11032-013-9917-x.
Gowda M., Das B., Makumbi D., et al. (2015) Genome-wide association and genomic prediction of resistance to maize lethal necrosis disease in tropical maize germplasm. Theoretical and Applied Genetics ;128(10):1957-1968.
Takagi H., Abe A., Yoshida K., et al. (2013) QTL-seq: rapid mapping of quantitative trait loci in rice by whole genome resequencing of DNA from two bulked populations. Plant Journal 74:174-183. DOI: 10.1111/tpj.12105.
Wangai A.W., Redinbaugh M.G., Kinyua Z.M., Miano D.W., Leley P.K., Kasina M., Mahuku G., Scheets K., Jeffers D. (2012) First report of Maize chlorotic mottle virus and maize lethal necrosis in Kenya. Plant Disease 96:1582-1583. DOI: 10.1094/pdis-06-12-0576-pdn.
Figure 1: Farmer fields in Kenya (left) and Babati, Tanzania (right) under natural
MLN disease pressure